The overall goal of the current application is to understand the neurobiological mechanisms that help confer pathological behaviors like enhanced negative affect following ethanol physical dependence. Recent studies suggest that intrinsic lateral/basolateral amygdala (BLA) GABAergic neurons tightly control the expression of negative emotions including those expressed during withdrawal following chronic ethanol exposure. Dopaminergic inputs from the ventral tegmentum/substantia nigra pars compacta have been shown to regulate this GABAergic system and disinhibit BLA principal neurons which drive the expression of anxiety-like behaviors. Based on strong preliminary evidence, our proposed experiments will test the central hypothesis that ethanol dependence dis-inhibits BLA output by dysregulating dopaminergic modulation of these GABAergic neurons. We will test our central hypothesis and accomplish our overall goal by utilizing a well- established rat model of chronic ethanol exposure and by integrating optogenetic, pre- and post-synaptic dopamine neurophysiology, and behavioral experimental approaches. The BLA has been extensively implicated as an important regulatory component of the neural circuitry controlling both anxiety-like behavior during withdrawal from chronic ethanol exposure as well as reward-seeking in drug-nave and -exposed animals. Specific Aim 1 will test our central hypothesis hypothesis by examining presynaptic dopamine function during withdrawal from ethanol dependence. We will directly measure DA release and reuptake in vitro by integrating in vitro fast-scan cyclic voltammetry with optogenetic control of DA release and chronic ethanol exposure. We hypothesize that, based on our previous publications, chronic ethanol exposure will differentially modulate basal or `tonic' DA levels and phasic DA release to ultimately enhance DA signaling. Specific Aim 2 will examine how postsynaptic DA receptor function is altered in the BLA using in vitro whole cell patch clamp electrophysiology with innovative optogenetic approaches to measure postsynaptic DA receptor signaling. Our working hypothesis is that ethanol physical dependence will increase postsynaptic D1- and D2-like DA receptor signaling such that DA-mediated inhibition of GABAergic function is up-regulated. Specific Aim 3 will place the detailed cellular effects of dependence on DA signaling within a whole-animal context by integrating optogenetic control of DA release with in vivo measures of BLA-dependent behaviors. Our working hypothesis is that dependence-related changes in dopamine neurotransmission and signaling ultimately control withdrawal-dependent anxiety-like behavior. The proposed work employs a unique and highly integrated experimental approach to provide unparalleled insight into the neurobiological mechanisms governing the negative reinforcing effects of chronic ethanol exposure. Ultimately, these studies will provide insight into potential cellular mechanisms governing abuse and relapse in human alcoholics.